Understanding the cellular mechanisms by which addictive substances modify the brain is a key target of research that is designed to provide a basis for development of treatment strategies for addicted patients. Our hypothesis is that a critical aspect of addiction is abberant glutamatergic neuronal plasticity involving mechanisms common to learning, such as LTP and LTD. The experiments proposed in this application will test this hypothesis by examining the role of transmembrane AMPA receptor regulatory proteins (TARPs) in both AMPA receptor trafficking and addiction-related plasticity in the nucleus accumbens (NAc), a brain region that plays a critical role in the development and persistence of addiction. I propose to investigate 1) the interactions between TARP isoforms and AMPA receptor subunits at the cellular level, 2) the role of TARP phosphorylation in AMPA receptor trafficking in the NAc, 3) the relationship between increases in AMPA receptor surface expression associated with enhanced drug-craving and TARP phosphorylation and surface expression in an animal model of addiction. TARPs have not previously been examine in the NAc. First, I will use immunocytochemical and quantitative co-immunoprecipitation techniques to determine the distribution of TARP isoforms and their association with AMPA receptor subunits in the NAc. Second, I will determine the effect of AMPA receptor synaptic incorporation on the synaptic incorporation of TARPs using immunocytochemistry and on TARP phosphorylation using Western blotting and isoelecrtric focusing in a novel NAc-prefrontal cortex co-culture system. Finally, I will examine the relationship between enhanced cue elicited drug-craving after withdrawal from cocaine self-administration and TARP phosphorylation and surface expression. To this end, NAc tissue from rats previously allowed to self-administer cocaine or saline will be collected after withdrawal and several biochemical techniques will be used to determine TARP phosphorylation, kinase activity, and surface expression of AMPA receptors and TARPs. Taken together, data from these experiments will provide essential and novel knowledge about the mechanisms underlying AMPA receptor trafficking in the NAc and will contribute to our understanding of glutamatergic plasticity associated with addiction. As many as 3 million Americans will use cocaine within their lifetimes, and addiction to cocaine (as well as other psychostimulants such as amphetamines) continues to be a health concern at the national level. The experiments proposed in this application will provide knowledge about the cellular mechanisms of AMPA receptor trafficking, and will also provide a link between alterations in the glutamatergic system and behaviors indicative of addiction, such as enhanced drug craving. [unreadable] [unreadable] [unreadable]